Effect of nitration on the physicochemical and kinetic features of wild-type and monotyrosine mutants of human respiratory cytochrome c.
The effect of tyrosine nitration on the physicochemical properties and reactivity of human respiratory cytochrome c has been extensively analyzed. A set of mutants, each bearing only one tyrosine out of the five present in the wild-type molecule, has been constructed in order to study the effect of each tyrosine nitration on the properties of the whole protein. Replacement of tyrosines by phenylalanines does not promote significant changes in the properties of the cytochrome. Nitration of wild-type cytochrome c promotes a drastic decrease (ca. 350 mV) in the midpoint redox potential, probably induced by nitration of both tyrosines 48 and 67. Nitration also promotes a significant decrease in the intrinsic reactivity of all the wild-type and mutant proteins. Nitration of mutant cytochromes and, in particular, of the wild-type protein significantly decreases their reactivity with cytochrome c oxidase, thereby suggesting that this alteration is due to an accumulative effect of different nitrations. The reactivity of mutants bearing tyrosine 67 and, to a lesser extent, tyrosine 74 is more affected by nitration, indicating that the change in reactivity of nitrated wild-type cytochrome c is mainly due to nitration of these tyrosine residues. Moreover, nitration of wild-type cytochrome c induces a significant loss in its ability to activate caspases because of the additive effect of nitration of several tyrosine groups, as inferred from the behavior of monotyrosine mutants.
Rodríguez-Roldán
Vicente
V
Instituto de Bioquímica Vegetal y Fotosíntesis, Centro de Investigaciones Científicas Isla de la Cartuja, Universidad de Sevilla and Consejo Superior de Investigaciones Científicas, Spain.
García-Heredia
José Manuel
JM
Navarro
José A
JA
De la Rosa
Miguel A
MA
Hervás
Manuel
M
D000818:Animals; D020169:Caspases / Q000378:metabolism; D019069:Cell Respiration; D045304:Cytochromes c / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003576:Electron Transport Complex IV / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005415:Flavins / Q000378:metabolism; D006736:Horses; D006801:Humans; D019169:Jurkat Cells; D007700:Kinetics; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009574:Nitro Compounds / Q000378:metabolism* / Q000494:pharmacology; D013057:Spectrum Analysis; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism*
D005415:Flavins; D050505:Mutant Proteins; D009574:Nitro Compounds; D014443:Tyrosine; D045304:Cytochromes c; D003576:Electron Transport Complex IV; D020169:Caspases